Transmembrane currents play a fundamental role in the activation and functioning of excitable tissues. In urinary bladder smooth muscle, depolarization, excitation-contraction, and repolarization are dependent upon the activation of transmembrane currents through voltage dependent ion channels. The current underlying repolarization in detrusor smooth muscle is carried through several ion channels, virtually all of which utilize potassium as the charge carrier. These include a transient, 4-aminopyridine sensitive current (Fujii K, Foster C D, Brading A F and Parekh A B. Potassium channel blockers and the effects of cromakalim on the smooth muscle of the guinea-pig bladder. Br J Pharmacol 99: 779–785, 1990), a delayed rectifier (Klöckner, U. and Isenberg, G. Calcium currents of cesium loaded isolated smooth muscle cells (urinary bladder of the guinea pig). Pflügers Arch 405: 340–348, 1985), an ATP-dependent current (Bonev A D and Nelson M T. ATP-sensitive potassium channels in smooth muscle cells from guinea pig urinary bladder. Am J Physiol 264(Cell Physiol 33): C1190–C1200, 1993; Trivedi S, Stetz S L, Potter-Lee L, McConville M, Li J H, Empfield J, Ohnmacht C J, Russell K, Brown F J, Trainor D A et al. K-channel opening activity of ZD6169 and its analogs: effect on 86Rb efflux and 3H-P1075 binding in bladder smooth muscle. Pharmacol 50: 388–397, 1994) and a charybdotoxin-sensitive current consistent with the large-conductance, calcium-dependent potassium current (BKCa) (Zografos P, Li J H and Kau S T. Comparison of the in vitro effects of K+ channel modulators on detrusor and portal vein strips from guinea pigs. Pharmacol 45: 216–230, 1992). Several of these channels have been the target of compounds and drugs aimed at modulating the physiology and functioning of smooth muscle and other tissues (Edwards, G. and Weston, A. H.: Pharmacology of the potassium channel openers. Cardiovasc Drugs and Ther 9: 185–193, 1995).
It has been suggested (Foster D C and Brading A F. The effect of potassium channel antagonists on the BRL 34915 activated potassium channel in guinea-pig bladder. Br J Pharmacol 92: 751, 1987) that a potassium channel opener (KCO) may be useful in the treatment of detrusor hyperactivity. An increase in potassium channel permeability would hyperpolarize the cell, bring the membrane potential further from the threshold for activation of calcium channels and reduce excitability (Brading A F. Ion channels and control of contractile activity in urinary bladder smooth muscle. Jap J Pharmacol 58 Suppl 2: 120P–127P, 1992). A number of potassium channel openers have shown activity in isolated tissues (Fujii et al., 1990; Malmgren A, Andersson K E, Andersson P O, Fovaeus M and Sjogren C. Effects of cromakalim (BRL 34915) and pinacidil on normal and hypertrophied rat detrusor in vitro. J Urol 143: 828–834, 1990; Grant T L and Zuzack J S. Effects of K+ channel blockers and cromakalim (BRL 34915) on the mechanical activity of guinea pig detrusor smooth muscle. J Pharmacol Exp Thera 269(3): 1158–1164, 1991) and efficacy in both experimental (Foster and Brading, 1987; Malmgren A, Andersson K E, Sjogren C and Andersson P O. Effects of pinacidil and cromakalim (BRL 34915) on bladder function in rats with detrusor instability. J Urol 142: 1134–1138, 1989; Wojdan A, Freeden C, Woods M, Norton W. Warga D, Spinelli W, Colatsky T, Antane M, Antane S, Butera J and Argentieri T M. Comparison of the potassium channel openers ZD6169, celikalim and WAY-133537 on isolated bladder tissue and in vivo bladder instability in the rat. J Pharmacol Exp Therap 289: 1410–1418, 1999) and clinical bladder instability (Nurse et al., 1991). However, because these compounds also activate channels in vascular smooth muscle (causing vasodilation), the clinical utility has been severely limited by hemodynamic side effects including hypotension and tachycardia.
It has been stated previously that retigabine (N-[2-amino-4-(4-fluorobenzylamino)-phenyl]carbamic acid ethyl ester) activates a member of the KCNQ family of potassium channel in the bladder which is most likely KCNQ2/3 and/or KCNQ3/5. (Wickenden A. D., Yu, W., Zou, A., Jegla, T., & Wagoner, P. K. Retigabine, a novel anti-convulsant, enhances activation of KCNQ2/Q3 potassium channels. Molec Pharmacol 58: 591–600 (2000); Wickenden, A. D., Zou, A., Wagoner, P. K., & Jela, T. Characterization of the KCNQ5/Q3 potassium channels expressed in mammalian cells. Br J Pharmacol 132: 381–384 (2001); Rundfeldt, C., Netzer, R. The novel anticonvulsant retigabine activates M-currents in Chinese hamster ovary-cells tranfected with human KCNQ2/3 subunits. Neuroscience Letters 282: 73–76 (2000); Main, M. J., Cryan, J. E., Dupere, J. R. B., Cox, B., Clare, J. J. & Burbidge, S. A. Modulation of KCNQ2/3 potassium channels by the novel anticonvulsant retigabine. Molec Pharm 58: 253–262 (2000)). The result is an inhibition of bladder smooth muscle contractility. In addition, recent data provides evidence for the existence of the KCNQ4 channel in human bladder smooth muscle. Current knowledge of KCNQ4 suggests that it may form a functional ion channel on its own (Søgaard S, Ljungstrøm T, Perersen K A, Olesen S P, Jensen, B S. KCNQ4 channels expressed in mammalian cells: functional characteristics and pharmacology. Am J Physiol 280: C859–C866, 2001), or that it may combine with KCNQ3 (Kubisch C. Schroeder B C. Friedrich T. Lutjohann B. El-Amraoui A. Marlin S. Petit C. Jentsch T J. KCNQ4, a novel potassium channel expressed in sensory outer hair cells, is mutated in dominant deafness. Cell 96(3):437–446, 1999). It is likely therefore, that retigabine's effects on bladder smooth muscle include activation of the KCNQ4 channel in addition to the channels formed by KCNQ2/3 and KCNQ3/5. Activation of this channel will hyperpolarize the bladder smooth muscle cells and, in doing so, relax the bladder. Since these KCNQ channels are not present in the cardiovascular system, retigabine and other molecules that activate these channels should be useful in the treatment of bladder instability without hemodynamic compromise.
M-currents have been shown to play an important functional role as determinants of cell excitability. Recent evidence indicates that the KCNQ potassium channel subunit form the molecular basis for M-current activity in a variety of tissues. From their initial report in peripheral sympathetic neurons the gene family has evolved to contain at least five major sub-units designated KCNQ1 though KCNQ5 (see reviews in Rogowski, M. A. KCNQ2/KCNQ3 K+ channels and the molecular pathogenesis of epilepsy: implications for therapy. TINS 23: 393–398, (2000); Jentsch, T. J. Neuronal KCNQ potassium channels: physiology and role in disease, Nature Rev, (2000)). These sub-units have been shown to co-assemble to form both heteromeric and homomeric functional ion channels. Recent reports indicate that both KCNQ2 and KCNQ5 can co-assemble with KCNQ3 (Tinel, N., Lauritzen, I., Chouabe, C., Lazdunski, M., Borsotto, M. The KCNQ2 potassium channel: splice variants, functional and developmental expression. Brain localization and comparison with KCNQ3. FEBS Letters. 438: 171–176 (1998); Yang, W., P., Levesque, P., C., Little, W., A., Conder, M., L., Ramakrishnan, P., Neubauer, M., G., Blanar, M., A. Functional expression of two KvLQT1-related potassium channels responsible for an inherited idiopathic epilepsy. J Biological Chemistry. 273:19419–19423 (1998); Wang, H. S., Pan, Z., Shi, W., Brown, B. S., Wymore, R. S., Cohen, I. S., Dixon, J. E. & McKinnon, D. KCNQ2 and KCNQ3 potassium channel subunits: molecular correlets of the M-channel. Science 282: 1890–1893, (1998); Lerche, C., Scherer, C. R., Seebohm, G., Derst, C., Wei, A. D., Busch, A. E., Steinmeyer, K. J Biologic Chem (2000); Schroeder, B., C., Hechenberger, M., Weinreich, F., Kubisch, C., Jentsch, T., J. KCNQ5, a novel potassium channel broadly expressed in brain, mediates M-type currents. [Journal Article] J Biological Chemistry. 275: 24089–24095 (2000)) to form a functional M-channel activatable by retigabine (Wickenden A. D., Yu, W., Zou, A., Jegla, T., & Wagoner, P. K. Retigabine, a novel anti-convulsant, enhances activation of KCNQ2/Q3 potassium channels. Molec Pharmacol 58: 591–600 (2000); Wickenden, A. D., Zou, A., Wagoner, P. K., & Jela, T. Characterization of the KCNQ5/Q3 potassium channels expressed in mammalian cells. Br J Pharmacol 132: 381–384 (2001); Rundfeldt, C., Netzer, R. The novel anticonvulsant retigabine activates M-currents in Chinese hamster ovary-cells transfected with human KCNQ2/3 subunits. Neuroscience Letters 282: 73–76 (2000); Main, M. J., Cryan, J. E., Dupere, J. R. B., Cox, B., Clare, J. J. & Burbidge, S. A. Modulation of KCNQ2/3 potassium channels by the novel anticonvulsant retigabine. Molec Pharm 58: 253–262 (2000)) and blocked by either acetylcholine (Adams, P., R., Brown, D., A., Constanti, A. M-currents and other potassium currents in bullfrog sympathetic neurones. J Physiology 330: 537–72(1982); Brown, D., A., Adams, P., R. Muscarinic suppression of a novel voltage-sensitive K+ current in a vertebrate neurone. Nature 283: 673–676(1980); Shapiro, M., S., Roche, J., P., Kaftan, E., J., Cruzblanca, H., Mackie, K., Hille, B. Reconstitution of muscarinic modulation of the KCNQ2/KCNQ3 K(+) channels that underlie the neuronal M current. J Neuroscience 20: 1710–1721 (2000)) linopirdine or XE-991 (10,10-bis(4-pyridinylmethyl)-9(10H)-anthracenone (Aiken, S. P., Lamp, B. J. Murphy, P. A. & Brown B. S. Reduction of spike frequency adaptation and blockade of M-current in rat CA1 pyramidal neurons by linopirdine (DuP 996) a neurotransmitter release enhancer. Br J Pharm 115: 1163–1168, (1995); Zaczek R. Chorvat R J. Saye J A. Pierdomenico M E. Maciag C M. Logue A R. Fisher B N. Rominger D H. Earl R A. Two new potent neurotransmitter release enhancers, 10,10-bis(4-pyridinylmethyl)-9(10H)-anthracenone and 10,10-bis(2-fluoro-4-pyridinylmethyl)-9(10H)-anthracenone: comparison to linopirdine. J Pharmacology & Exp Therap 285: 724–730 (1998). The parasympathetic neurotransmitter acetylcholine (Ach) is known to produce several physiological responses in bladder smooth muscle. The net result of Ach exposure is a contraction of the smooth muscle mainly through the mobilization of transmembrane and intracellular calcium stores (Hashitani H. Bramich N J. Hirst G D. Mechanisms of excitatory neuromuscular transmission in the guinea-pig urinary bladder. Journal of Physiology 524: 565–579 (2000)). The role that Ach plays in modulating the cell transmembrane potential, however, is more complex. Pathways for both hyperpolarization and depolarization are present with muscarinic stimulation of bladder smooth muscle. Hyperpolarization may be associated with a mechanism that involves calcium sparks and activation of calcium-dependent potassium currents (Herrera G M. Heppner T J. Nelson M T. Voltage dependence of the coupling of Ca(2+) sparks to BK(Ca) channels in urinary bladder smooth muscle. American Journal of Physiology—Cell Physiology 280: C481–490 (2001)).
Furthermore, there is a need to develop methods of selecting compounds useful in the treatment bladder instability and related urologic or bladder conditions. The present invention meets this need and includes methods of treatment of bladder instability and related urologic and bladder conditions.